CN103060238A - Micro-ecological water purifying agent for aquatic products, and preparation method and application thereof - Google Patents
Micro-ecological water purifying agent for aquatic products, and preparation method and application thereof Download PDFInfo
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Abstract
The invention relates to a micro-ecological water quality adjusting agent for aquatic product culture, and a preparation method and an application thereof. The agent comprises 1 part of Bacillus cereus, 1 part of Bacillus laterosporus and 1 part of Saccharomyces cerevisiae. The preparation method comprises the following steps: respectively carrying out submerged liquid fermentation of the Bacillus cereus, the Bacillus laterosporus and the Saccharomyces cerevisiae, and uniformly mixing according to a volume ratio of 1:1:1. The micro-ecological water purifying agent is utilized to improve water in the invention, so the dissolved oxygen (DO) in water can be improved, and the contents of ammonia nitrogen, nitrites and COD in water can be improved, thereby the water quality is improved, and the healthy growth of fish and shrimps is promoted; and the microbial community structure in water is changed, and the pathogen growth is inhibited, so the generation of diseases of aquatic product animals comprising the fish, the shrimps and the like is effectively prevented, and the survival rates of the aquatic product animals comprising the fish, the shrimps and the like are improved.
Description
Technical field
The present invention relates to a kind of microecological water matter conditioning agent and methods for making and using same thereof for used for aquiculture, belong to microbial preparation technical field and biological environmental production field.
Background technology
Along with developing rapidly of China's culture fishery, some restriction culture fisheries problem healthy, sustainable development also highlights day by day.One, the discharging of a large amount of trade effluents, sanitary sewage has caused disastrous effect to aquaculture water; Its two, breeding production lacks comprehensive planning and effectively management, blindly the unordered pond of making increases net cage on a large scale, the breeding way of the high production that covets has aggravated the deterioration of water quality especially; Its three, the abuse of the residual bait that unreasonable bait throwing in produces, ight soil and fishing medicine is seriously damaged the water body microecological balance, has caused direct or indirect negative impact for the aquatic products aquaculture.In view of above reason, aquiculture disease frequent occurrence, fishery products lose intrinsic delicate flavour and body eutrophication etc. in recent years, and culture fishery has been caused serious financial loss.Therefore taking biological means and high-tech means pollution control of water, promote aquaculture to develop in a healthy way, guarantee the benign cycle of water ecological setting, has been instant task.
Summary of the invention
The present invention is directed to Aquaculture Status, take full advantage of the physiological function of microorganism and the biological restoration modeling effort that the modifiability characteristics are polluted aquaculture water, by separating, screen, tame efficient degrading bacterial strain, and utilize a kind of complex microorganism preparations of having complementary functions property structure of each bacterial strain, and provide its methods for making and using same.The concentration of the chemical oxygen demand (COD) in the reduction water body and ammonia nitrogen, nitrite is improved the cultivation water environment, improves the immunity of organisms of cultivation object, and the generation of minimizing disease makes the aquaculture cycle development.
A kind of aquatic products preparation method of microbial water purified agents, by bacillus cereus (Bacillus cereus) CGMCC No.4918, bacillus laterosporus (bacillus laterosporus) CGMCC No.6885 and yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) CGMCC No.6884, it is formulated that the zymocyte liquid of these 3 kinds of bacterium is pressed the volume ratio of 1:1:1.
The preparation of the zymocyte liquid of described bacillus cereus is as follows successively:
Fresh slant strains preparation, its condition is: LB substratum, 30 ℃, 24h;
Table concentration seed culture, its condition is: 30 ℃ ~ 32 ℃ of pH7.2 ~ 7.5, temperature, rotating speed 200r/min ~ 220r/min cultivates 10 ~ 12h; Substratum consist of peptone 10g, NaCl5g, extractum carnis 3g, distilled water is settled to 1L;
Liquid fermenting, its condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 200r/min ~ 220r/min, initial pH7.0 ~ 7.2, ventilation ratio is 1.1:1 ~ 1.2:1, and table concentration seed culture liquid inoculum size is 3% ~ 4%, 30 ℃ ~ 32 ℃ temperature bottom fermentation 24h ~ 28h of culture volume; Substratum consists of: W-Gum 1.8% ~ 2.0%, industrial peptone 1.0% ~ 1.2%, MnSO
40.02%, CaCl
20.02% ~ 0.03%, K
2HPO
40.012%, KH
2PO
40.025%, all the other are water; Each fermentation gemma number is 1.8 * 10
10Cfu/mL ~ 2.5 * 10
10Between the cfu/mL.
The preparation of the zymocyte liquid of described bacillus laterosporus is as follows successively:
Fresh slant strains preparation, its condition is: LB substratum, 35 ℃, 28h;
Table concentration seed culture, its condition is: 32 ℃ ~ 35 ℃ of pH7.0 ~ 7.2, temperature, rotating speed 180r/min ~ 200r/min cultivates 14h ~ 16h; Consisting of of substratum: peptone 10g, NaCl5g, extractum carnis 3g, distilled water is settled to 1L;
Liquid fermenting, its condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 180r/min ~ 200r/min, initial pH7.0 ~ 7.2, ventilation ratio 1:1 ~ 1.1:1, table concentration seed culture liquid inoculum size is 3% ~ 4%, 32 ℃ ~ 35 ℃ temperature bottom fermentation 28h ~ 36h of culture volume; Substratum consists of: glucose 2.0% ~ 2.2%, yeast extract paste 0.9% ~ 1.1%, MnSO
40.02%, CaCl
20.02% ~ 0.03%, KH
2PO
40.01%, MgSO
40.005%, all the other are water; Each fermentation gemma number is 8 * 10
9Cfu/mL ~ 1.02 * 10
10Between the cfu/mL.
The preparation of the zymocyte liquid of described yeast saccharomyces cerevisiae is as follows successively:
Fresh slant strains preparation, its condition is: PDA substratum, 28 ℃, 36h ~ 48h;
Table concentration seed culture, its condition is: 26 ℃ ~ 28 ℃ of pH nature, temperature, rotating speed 180r/min ~ 200r/min cultivates 24h ~ 28h; Substratum consist of glucose 20g, yeast extract paste 10g, peptone 10g, distilled water is settled to 1L;
Liquid fermenting, its condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 180r/min ~ 200r/min, initial pH6.0 ~ 6.5, ventilation ratio is 0.9:1～1:1, and table concentration seed culture liquid inoculum size is 4% ~ 5%, 26 ℃ ~ 28 ℃ temperature bottom fermentation 28h ~ 36h of culture volume; Substratum consists of sucrose 2% ~ 2.5%, ammonium sulfate 1% ~ 1.5%, and calcium chloride 0.5% ~ 0.8%, vitamin H 0.01%, potassium primary phosphate 0.01%, sodium-chlor 0.01%, sal epsom 0.03% ~ 0.06%, all the other are water; The zymophyte number of yeast saccharomyces cerevisiae reaches 7.4 * 10
8Cfu/mL ~ 9.5 * 10
8Cfu/mL.
With the zymocyte liquids of three kinds of bacterium 1:1:1 stirring and evenly mixing by volume, stirring velocity 50r/min ~ 80r/min, churning time 10min ~ 30min makes the aquatic products microbial water purified agents, and its finished product viable count is not less than 9 * 10
9Cfu/mL.
A kind of aquatic products microbial water purified agents is to be prepared from by above-mentioned method.
Above-mentioned aquatic products is with the application method of microbial water purified agents: put in the water body, be used for the purifying aquatic product water body.
A kind of is bacillus laterosporus (Bacillus laterosporus) for purifying aquatic product with the bacterium of water body, deposit number CGMCC No.6885.
A kind of is yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) for purifying aquatic product with the fungi of water body, deposit number CGMCC No.6884.
Yeast saccharomyces cerevisiae of the present invention (Saccharomyces cerevisiae) CGMCC No.6884:
Bacterium colony and morphological features
Bacterial strain CGMCC No.6884 colonial morphology is oyster white, and is opaque, moistening, neat in edge.Thalli morphology is circular, produces thecaspore, forms original mycelia or becomes the short chain shape, without ballistospore throw out (Fig. 1, Fig. 2).
Physio-biochemical characteristics
The physiological and biochemical property of bacterial strain CGMCC No.6884 sees Table 1 ~ 4.
Table 1 fermenting carbohydrate
Table 2 assimilation carbon source
Table 3 assimilation nitrogenous source
Other physiological and biochemical tests of table 4
Ammonia nitrogen in yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) the CGMCC No.6884 energy efficient degradation water body, when the ammonia nitrogen starting point concentration is 212mg/L, inoculum size 3%, degradation rate is 91.4%; Fermented liquid diastatic activity 28.4MMU, protease activity 215U; Inhibition zone size to intestinal bacteria (Escherichia coli), staphylococcus aureus (Staphyloccocus aureus), Salmonellas (Salmonella sp.), Pseudomonas fluorescens (Pseudomonas fluorescens) and Aeromonas hydrophila (Aeromonas hydrophila) is respectively 21.3mm, 16.7mm, 23.8mm, 17.9mm, 20.7mm.
Bacillus cereus (Bacillus cereus) CGMCC No.4918
Ne ar and colony characteristics: bacterial strain CGMCC No.4918 bacterium colony is larger, circle, oyster white, neat in edge, smooth, the thickness of bacterium colony surface wettability.It is shaft-like that thalline is, terminal blunt circle, and (0.8 ~ 1.2) μ m * (1.8 ~ 3.8) μ m produces gemma, and atrichia is without pod membrane (seeing Fig. 3, Fig. 4).Gram-positive.
The physiological and biochemical property of bacterial strain CGMCC No.4918 is: Starch Hydrolysis, Citrate trianion utilization, nitrate reduction, catalase, litmus milk test, indole test, V-P test, methyl red test, gelatin hydrolysis test, the 0.001% N,O-Diacetylmuramidase resistant proof positive; Urea test, hydrogen sulfide production test feminine gender; Utilize sucrose, glucose, fructose, maltose, lactose to produce acid, can not utilize semi-lactosi, N.F,USP MANNITOL to produce acid.Growth temperature is 5 ℃ ~ 45 ℃.
Bacillus cereus (Bacillus cereus) CGMCC No.4918, the nitrite in the energy efficient degradation water body, when the nitrite starting point concentration is 10mg/mL, inoculum size 3%, degradation rate is 88.8%; Fermented liquid diastatic activity 35.5MMU, protease activity 268U; Inhibition zone size to intestinal bacteria, staphylococcus aureus, Salmonellas, Pseudomonas fluorescens and Aeromonas hydrophila is respectively 23.6mm, 16.9mm, 20.4mm, 15.9mm, 15.7mm.
Bacillus laterosporus of the present invention (Bacillus laterosporus) CGMCC No.6885
Ne ar and colony characteristics: bacterial strain C5 bacterium colony is larger, circle, oyster white, neat in edge, smooth, the thickness of bacterium colony surface wettability.It is shaft-like that thalline is, and (0.5 ~ 0.8) μ m * (2.5 ~ 4.7) μ m has paddle shape side spore, and gemma is oval.Gram-positive.(seeing Fig. 5, Fig. 6).
Physio-biochemical characteristics: Starch Hydrolysis, nitrate reduction, hydrogen sulfide production test, gelatin hydrolysis test, catalase, the urea test positive, can tolerate 7% NaCl and 0.001% N,O-Diacetylmuramidase, on the substratum of pH5-7 and nitrogen-free agar, all can grow, utilize sucrose, glucose, N.F,USP MANNITOL; Can not utilize wood sugar, lactose, pectinose.
Bacillus laterosporus (CGMCC No.6885), the COD in the energy efficient degradation water body, when the COD starting point concentration is 456.2mg/L, inoculum size 3%, degradation rate reaches 94.1%.Fermented liquid diastatic activity 31.7MMU, protease activity 323U; Inhibition zone size to intestinal bacteria, staphylococcus aureus, Salmonellas, Pseudomonas fluorescens and Aeromonas hydrophila is respectively 18.8mm, 16.7mm, 16.9mm, 12.9mm, 21.3mm.
Bacillus cereus of the present invention (Bacillus cereus) CGMCC No.4918 has carried out the patent preservation on May 30th, 2011 at China Committee for Culture Collection of Microorganisms common micro-organisms center; Biological depositary institution address is No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica.
Bacillus laterosporus of the present invention (bacillus laterosporus) CGMCC No.6885 has carried out the patent preservation on November 26th, 2012 at China Committee for Culture Collection of Microorganisms common micro-organisms center; Biological depositary institution address is the Datun Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica.
Yeast saccharomyces cerevisiae of the present invention (Saccharomyces cerevisiae) CGMCC No.6884 has carried out the patent preservation on November 26th, 2012 at China Committee for Culture Collection of Microorganisms common micro-organisms center; Biological depositary institution address is the Datun Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica.
Above-mentioned aquatic products reduces its content in water body with the pollutents such as ammonia nitrogen in the water body and nitrite that can directly receive, digest, degrade of the beneficial microorganism in the microbial water purified agents, improves Water quality; Beneficial microorganism produces various meta-bolitess such as microbiotic, enzyme etc. in the growth and breeding process, the growth of energy establishment pathogenic micro-organism reduces pathogenic agent, reduces the generation of Animal diseases; Beneficial microorganism can be kept the eubiosis of animal intestinal microenvironment, improves efficiency of feed utilization, improves animal immunizing power, promotes growth of animal.By strong, stable, the efficient microorganism strains of screening field planting power, according to the bacterial strain effect of having complementary functions, complex micro organism fungicide is made in active combination, rationally use, realize the cultivation of water body animal health cleaning, effectively improve aquaculture to the impact of ecotope, make aquifer cultivation stablize sustainable development.
Description of drawings:
Fig. 1: the colonial morphology figure of yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) CGMCC No.6884;
Fig. 2: the morphological features figure of yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) CGMCC No.6884;
Fig. 3: the colonial morphology figure of bacillus cereus (Bacillus cereus) CGMCC No.4918;
Fig. 4: the morphological features figure of bacillus cereus (Bacillus cereus) CGMCC No.4918;
Fig. 5: the colonial morphology figure of bacillus laterosporus (Bacillus laterosporus) CGMCC No.6885;
Fig. 6: the morphological features figure of bacillus laterosporus (Bacillus laterosporus) CGMCC No.6885;
Fig. 7: different probioticses are on the impact of simulation water body COD;
Fig. 8: different probioticses are on the impact of simulation water body ammonia nitrogen;
Fig. 9: different probioticses are on the impact of simulation water nitrite;
Figure 10: different probioticses are on the impact of simulation Dissolved Oxygen in Water;
Figure 11: different probioticses are on the impact of simulation water pH value.
Embodiment
Be intended to further specify the present invention below in conjunction with embodiment, and unrestricted the present invention.
Embodiment 1:
The aquatic products preparation of microbial water purified agents
1. the preparation of bacillus cereus (Bacillus cereus) CGMCC No.4918 bacterium liquid
Fresh slant strains preparation, its condition is: LB substratum, 30 ℃, 24h;
The picking slant strains is carried out the shaking table seed liquor and is cultivated: 30 ℃ ~ 32 ℃ of pH7.2 ~ 7.5, temperature, and rotating speed 200r/min ~ 220r/min cultivates 10 ~ 12h; Substratum consist of peptone 10g, NaCl5g, extractum carnis 3g, distilled water is settled to 1L;
Liquid fermentation condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 200r/min ~ 220r/min, initial pH7.0 ~ 7.2, ventilation ratio is 1.1:1 ~ 1.2:1, table concentration seed culture liquid inoculum size is 3% ~ 4%, 30 ℃ ~ 32 ℃ temperature bottom fermentation 24h ~ 28h of culture volume; Substratum consists of: W-Gum 1.8% ~ 2.0%, industrial peptone 1.0% ~ 1.2%, MnSO
40.02%, CaCl
20.02% ~ 0.03%, K
2HPO
40.012%, KH
2PO
40.025%, all the other are water; Each fermentation gemma number is 1.8 * 10
10Cfu/mL ~ 2.5 * 10
10Between the cfu/mL;
2. the preparation of bacillus laterosporus (Bacillus laterosporus) CGMCC No.6885 bacterium liquid
Fresh slant strains preparation, its condition is: LB substratum, 35 ℃, 28h;
The picking slant strains is carried out the shaking table seed liquor and is cultivated: 32 ℃ ~ 35 ℃ of pH7.0 ~ 7.2, temperature, and rotating speed 180r/min ~ 200r/min cultivates 14h ~ 16h; Consisting of of substratum: peptone 10g, NaCl5g, extractum carnis 3g, distilled water is settled to 1L;
Liquid fermentation condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 180r/min ~ 200r/min, initial pH7.0 ~ 7.2, ventilation ratio 1:1 ~ 1.1:1, table concentration seed culture liquid inoculum size is 3% ~ 4%, 32 ℃ ~ 35 ℃ temperature bottom fermentation 28h ~ 36h of culture volume; Substratum consists of: glucose 2.0% ~ 2.2%, yeast extract paste 0.9% ~ 1.1%, MnSO
40.02%, CaCl
20.02% ~ 0.03%, KH
2PO
40.01%, MgSO
40.005%, all the other are water; Each fermentation gemma number is 8 * 10
9Cfu/mL ~ 1.02 * 10
10Between the cfu/mL;
3. the preparation of yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) CGMCC No.6884 bacterium liquid
Fresh slant strains preparation, its condition is: PDA substratum, 28 ℃, 36h ~ 48h;
The picking slant strains is carried out the shaking table seed liquor and is cultivated: 26 ℃ ~ 28 ℃ of pH nature, temperature, and rotating speed 180r/min ~ 200r/min cultivates 24h ~ 28h; Substratum consist of glucose 20g, yeast extract paste 10g, peptone 10g, distilled water is settled to 1L;
Liquid fermentation condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 180r/min ~ 200r/min, initial pH6.0 ~ 6.5, ventilation ratio is 0.9:1～1:1, table concentration seed culture liquid inoculum size is 4% ~ 5%, 26 ℃ ~ 28 ℃ temperature bottom fermentation 28h ~ 36h of culture volume; Substratum consists of sucrose 2% ~ 2.5%, ammonium sulfate 1% ~ 1.5%, and calcium chloride 0.5% ~ 0.8%, vitamin H 0.01%, potassium primary phosphate 0.01%, sodium-chlor 0.01%, sal epsom 0.03% ~ 0.06%, all the other are water; The zymophyte number of yeast saccharomyces cerevisiae reaches 7.4 * 10
8Cfu/mL ~ 9.5 * 10
8Cfu/mL;
4. aquatic products is used the composite of microbial water purified agents
With the above-mentioned 3 kinds of bacterium liquid through being up to the standards by volume 1:1:1 move into stirring and evenly mixing in the sterilized storage tank, stirring velocity 50r/min ~ 80r/min, then churning time 10min ~ 30min carries out canned between aseptic canning.The finished product viable count that makes is not less than 9 * 10
9Cfu/mL.
Embodiment 2:
Aquatic products probiotics safety evaluation
Probiotics is applied in the aquaculture, the most important condition to aquatic animal is safety, toxicological harmless effect, in order to ensure application safety, the present invention is from hemolytic test, acute toxicity test in mice and three aspects of zebra fish acute toxicity test, inquire into the security of estimating said preparation, for its further Application and Development provides the safety experiment data.
The hemolytic test-results shows that the hemolysis rate of each bacterial strain and probiotics all is lower than 5%, shows that each bacterial strain and probiotics can not cause erythrocytic obvious aggegation and haemolysis, and security is better.The acute toxicity test in mice result shows the oral mld (LD of probiotics
50) greater than 75000mg/kg Mice Body quality.The zebra fish the acute toxicity tests shows that probiotics is to the 96h-LD of zebra fish
50Greater than 10000mg/L.Above test-results shows this probiotics safety non-toxic, can be used for aquaculture water.
Embodiment 3:
The aquatic products application of microbial water purified agents
At laboratory preparation breeding wastewater, initial condition of water quality is COD:162.75mg/L; Ammonia nitrogen: 12.05mg/L; Nitrite: 5.33mg/L; Dissolved oxygen: 4.62mg/L; PH:7.53.Breeding wastewater places respectively the 1000mL Erlenmeyer flask, test divides 5 groups, test group 1 is pressed addition 1mL/L and is added bacillus cereus CGMCC No.4918 bacterium liquid, test group 2 is pressed addition 1mL/L and is added bacillus laterosporus CGMCC No.6885 bacterium liquid, test group 3 is pressed addition 1mL/L and is added yeast saccharomyces cerevisiae CGMCC No.6884 bacterium liquid, test group 4 is pressed addition 1mL/L and is added aquatic products microbial water purified agents of the present invention, and control group adds 1000mL waste water, does not add any bacterium liquid.Every 3d measures COD, ammonia nitrogen, nitrite, the content of dissolved oxygen and the variation of pH, test period 15d.Every group three parallel, and testing data is averaged.The mensuration of ammonia nitrogen adopts the nessler reagent spectrophotometry; The mensuration of nitrite adopts the hydrochloride naphthodiamide spectrophotometry; COD adopts potassium dichromate process to measure; Adopt respectively dissolved oxygen meter and pH meter to measure DO value and pH value.
Test-results shows that behind the test 15d, the concentration of the COD of each test group, ammonia nitrogen, nitric nitrogen has reduction in various degree, and dissolved oxygen has lifting in various degree, and wherein the improved effect of 4 pairs of water quality of test group is best.Behind the test 15d, test group 4COD final concentration is 11.93mg/L, and degradation rate reaches 92.3%; The ammonia nitrogen final concentration is 1.92mg/L, and degradation rate reaches 84.1%; The nitrite final concentration is 0.63mg/L, and degradation rate reaches 88.1%; Dissolved oxygen concentration is 6.17mg/L, has promoted 33.5%.And control group COD final concentration is 153.1mg/L, degradation rate only 5.9%; The ammonia nitrogen final concentration is 10.68mg/L, degradation rate only 11.3%; Nitrite final concentration 4.92mg/L, degradation rate only 7.6%; Dissolved oxygen concentration almost remains unchanged.The action effect of the more single bacterial strain probiotics of composite bacteria preparation is better, may be between each dominant bacteria synergy to be arranged, and tool is complementary on physiological function and genetic information, thereby the higher biological activity of tool.The pH value of each test group and control group in the 15d of test, is simulated the pH value of water body between 7.18 ~ 7.68, the cultivation of the suitable most of fishery products of this pH value without significant difference.
Embodiment 4:
Aquatic products is with the impact of microbial water purified agents on fishpond water quality and Growth of Grass Carps Ctenopharyngodon Idellus performance
Test in place is carried out in the thousand mu of plants in Huang Gai lake, Linxiang, Yueyang, and experimental animal is grass carp.Test is established a control group, a test group, every group of 3 repetitions.The test pool only adds microbial water purified agents (addition is 1mL/L), does not add microbiotic and other drug.The contrast pool does not add microbial water purified agents, according to conventional cultivation.Plant is cement pit (3.0m * 2.5m * 1.2m), totally 6 mouthfuls, average depth 0.7m; Fish 12 tails, totally 72 tails are put in every pond.Experiment periods is thrown something and fed 2 every day; 8:00,17:00, throwing the rate of raising is fish body weight about 2%, the basically identical throwing of each group maintenance is raised water gaging and is put down, and according to growth, the situation of ingesting adjustment.Culturing pool 24h uninterruptedly inflates.Water temperature is 24 ~ 28 ℃ during the cultivation, pH7.6 ± 0.5, dissolved oxygen 6.7 ~ 7.4mg/L.Every 30d gets the microbial water purified agents full pool of evenly splashing, and then COD, ammonia nitrogen and the nitrite concentration on each pool of experimental determination taken back in every 10d sampling.Every pool is established 3 sampling spots (get 3 some A, B, C in all directions in every mouthful of fish pond direction, wherein the B point is the central point in fish pond, and A and C point all are positioned at the B point to the point midway at diagonal angle) and is averaged.Test and counted 0d, test period 60d the same day.
Detection method is referring to embodiment 3, and all samples is measured 3 times, averages.After cultivation finished, the hungry 24h of fish body claimed every pond fish gross weight, calculates rate of body weight gain, feed coefficient, surviving rate; The close grass carp of 3 urosome quality is got in every pond at random, measures its height, long, the body weight of body, claims after dissecting that internal organ are heavy, liver is heavy, calculates coefficient of condition, internal organ ratio, liver proportion, the long height ratio of body.
This test shows that the microbial water purified agents of this laboratory development has good purifying water effect: good to the ammonia nitrogen in the aquaculture water, nitrite and COD degradation effect.After the off-test, ammonia nitrogen is reduced to 0.16mg/L from starting point concentration 2.82mg/L, and nitrite is reduced to 0.12mg/L from starting point concentration 0.52mg/L, and COD reduces to 9.1mg/L from starting point concentration 63.4mg/L, dissolved oxygen is promoted to 5.81mg/L from starting point concentration 4.25mg/L, and the pH value stabilization is about 7.2.And microbial water purified agents has improved the growth performance of cultivated animals: the trend that the test group of the microbial water purified agents of splashing in the water body is compared with control group and is improved rate of body weight gain, reduces feed coefficient; Each organizes grass carp aspect the body index, comprises the long height ratio of body, internal organ ratio, liver anharmonic ratio and coefficient of condition, all without significant difference (P〉0.05).
Claims (7)
1. an aquatic products is with the preparation method of microbial water purified agents, it is characterized in that, by bacillus cereus (Bacillus cereus) CGMCC No.4918, bacillus laterosporus (bacillus laterosporus) CGMCC No.6885 and yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) CGMCC No.6884, it is formulated that the zymocyte liquid of these 3 kinds of bacterium is pressed the volume ratio of 1:1:1.
2. aquatic products according to claim 1 is characterized in that with the preparation method of microbial water purified agents,
1) preparation of bacillus cereus fermentation bacterium liquid may further comprise the steps successively
Fresh slant strains preparation, its condition is: LB substratum, 30 ℃, 24h;
Table concentration seed culture, its condition is: 30 ℃ ~ 32 ℃ of pH7.2 ~ 7.5, temperature, rotating speed 200r/min ~ 220r/min cultivates 10 ~ 12h; Substratum consist of peptone 10g, NaCl5g, extractum carnis 3g, distilled water is settled to 1L;
Liquid fermenting, its condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 200r/min ~ 220r/min, initial pH7.0 ~ 7.2, ventilation ratio is 1.1:1 ~ 1.2:1, and table concentration seed culture liquid inoculum size is 3% ~ 4%, 30 ℃ ~ 32 ℃ temperature bottom fermentation 24h ~ 28h of culture volume; Substratum consists of: W-Gum 1.8% ~ 2.0%, industrial peptone 1.0% ~ 1.2%, MnSO
40.02%, CaCl
20.02% ~ 0.03%, K
2HPO
40.012%, KH
2PO
40.025%, all the other are water; Each fermentation gemma number is 1.8 * 10
10Cfu/mL ~ 2.5 * 10
10Between the cfu/mL;
2) preparation of bacillus laterosporus zymocyte liquid may further comprise the steps successively:
Fresh slant strains preparation, its condition is: LB substratum, 35 ℃, 28h;
Table concentration seed culture, its condition is: 32 ℃ ~ 35 ℃ of pH7.0 ~ 7.2, temperature, rotating speed 180r/min ~ 200r/min cultivates 14h ~ 16h; Consisting of of substratum: peptone 10g, NaCl5g, extractum carnis 3g, distilled water is settled to 1L;
Liquid fermenting, its condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 180r/min ~ 200r/min, initial pH7.0 ~ 7.2, ventilation ratio 1:1 ~ 1.1:1, table concentration seed culture liquid inoculum size is 3% ~ 4%, 32 ℃ ~ 35 ℃ temperature bottom fermentation 28h ~ 36h of culture volume; Substratum consists of: glucose 2.0% ~ 2.2%, yeast extract paste 0.9% ~ 1.1%, MnSO
40.02%, CaCl
20.02% ~ 0.03%, KH
2PO
40.01%, MgSO
40.005%, all the other are water; Each fermentation gemma number is 8 * 10
9Cfu/mL ~ 1.02 * 10
10Between the cfu/mL;
3) preparation of fermentation by saccharomyces cerevisiae bacterium liquid may further comprise the steps successively:
Fresh slant strains preparation, its condition is: PDA substratum, 28 ℃, 36h ~ 48h;
Table concentration seed culture, its condition is: 26 ℃ ~ 28 ℃ of pH nature, temperature, rotating speed 180r/min ~ 200r/min cultivates 24h ~ 28h; Substratum consist of glucose 20g, yeast extract paste 10g, peptone 10g, distilled water is settled to 1L;
Liquid fermenting, its condition is: 20L fermentation tank culture medium loading amount is 12L, stirring velocity 180r/min ~ 200r/min, initial pH6.0 ~ 6.5, ventilation ratio is 0.9:1～1:1, and table concentration seed culture liquid inoculum size is 4% ~ 5%, 26 ℃ ~ 28 ℃ temperature bottom fermentation 28h ~ 36h of culture volume; Substratum consists of sucrose 2% ~ 2.5%, ammonium sulfate 1% ~ 1.5%, and calcium chloride 0.5% ~ 0.8%, vitamin H 0.01%, potassium primary phosphate 0.01%, sodium-chlor 0.01%, sal epsom 0.03% ~ 0.06%, all the other are water; The zymophyte number of yeast saccharomyces cerevisiae reaches 7.4 * 10
8Cfu/mL ~ 9.5 * 10
8Cfu/mL.
3. aquatic products according to claim 2 is characterized in that with the preparation method of microbial water purified agents,
With the zymocyte liquids of three kinds of bacterium 1:1:1 stirring and evenly mixing by volume, stirring velocity 50r/min ~ 80r/min, churning time 10min ~ 30min makes the aquatic products microbial water purified agents, and its finished product viable count is not less than 9 * 10
9Cfu/mL.
4. an aquatic products microbial water purified agents is characterized in that, is prepared from by each described method of claim 1-3.
5. aquatic products claimed in claim 4 is characterized in that with the application method of microbial water purified agents, puts in the water body, is used for the purifying aquatic product water body.
6. one kind is used for purifying aquatic product with the bacterium of water body, it is characterized in that described bacterium is bacillus laterosporus (Bacillus laterosporus), deposit number CGMCC No.6885.
7. one kind is used for purifying aquatic product with the fungi of water body, it is characterized in that described fungi is yeast saccharomyces cerevisiae (Saccharomyces cerevisiae), deposit number CGMCC No.6884.
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| CN110980942A (en) * | 2019-12-10 | 2020-04-10 | 浙江永续环境工程有限公司 | Anaerobic biological agent and anaerobic treatment method using same |
| CN115806349A (en) * | 2022-12-15 | 2023-03-17 | 鹤山市新的生物制品有限公司 | Method for increasing dissolved oxygen content of aquaculture water and special culture medium |
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